Hek293t cell strain having high dispersibility and screening method therefor

Abstract

Provided are a HEK293T cell strain having high dispersibility and a screening method therefor. Specifically, the present application relates to a method for screening a HEK293T cell strain suitable for serum-free suspension culture, a cell strain screened by using the method, and a method for producing a viral vector by using the cell strain.

1 . A novel human embryonic kidney HEK293T cell strain, deposited at the China General Microbiological Culture Collection Center with a deposit number of CGMCC NO.: 21100. 
     
     
         2 . A method for screening a HEK293T cell strain suitable for serum-free suspension culture, comprising:
 i) taking a HEK293T cell adherently cultured in a serum-containing medium, and placing the cell in a serum-free medium for suspension culture; and   ii) selecting a monoclonal cell strain suitable for serum-free suspension culture.   
     
     
         3 . The method of  claim 2 , wherein the serum-containing medium in step i) contains 10% serum. 
     
     
         4 . The method of  claim 2 , wherein the suspension culture in step i) includes shaking culture at 37° C. and 8% CO 2 . 
     
     
         5 . The method of  claim 2 , wherein the suspension culture in step i) includes cell passaging. 
     
     
         6 . The method of  claim 2 , wherein step ii) includes selecting a non-clumping monoclonal cell strain suitable for serum-free suspension culture. 
     
     
         7 . The method of  claim 2  or  6 , wherein step ii) includes culturing the cell in a semi-solid medium, selecting a single cell from the semi-solid medium, and performing suspension culture on the cell. 
     
     
         8 . The method of  claim 7 , wherein the suspension culture of the selected cell in step ii) includes shaking culture at 37° C. and 8% CO 2 . 
     
     
         9 . The method of  claim 7 , wherein the suspension culture of the selected cell in step ii) includes cell passaging. 
     
     
         10 . The method of  claim 7 , wherein the semi-solid medium is a methylcellulose-based medium supplemented with a screening medium. 
     
     
         11 . The method of  claim 2 , wherein the serum-free medium is selected from LV-MAX production medium, Expi293 expression medium and OPM-293 CD03 medium. 
     
     
         12 . The method of  claim 2 , wherein the method is carried out without the addition of an anti-clumping agent. 
     
     
         13 . A cell strain screened by using the method of  claim 2 . 
     
     
         14 . The cell strain of  claim 13 , wherein the cell strain does not clump in a liquid medium. 
     
     
         15 . Use of a medium selected from LV-MAX production medium, Expi293 expression medium and OPM-293 CD03 medium in screening a non-clumping HEK293T cell strain suitable for suspension culture. 
     
     
         16 . A method for preparing a virus vector using the cell strain of  claim 1 , comprising:
 i) performing suspension culture on the cell strain;   ii) introducing a virus vector expression system into the cell strain;   iii) culturing the cell strain under a condition favorable for the production of the viral vector; and   vi) collecting the viral vector.   
     
     
         17 . (canceled) 
     
     
         18 . The method of  claim 16 , wherein the viral vector expression system comprises a packaging nucleic acid vector and an envelope nucleic acid vector. 
     
     
         19 . The method of  claim 16 , wherein the viral vector expression system further comprises an accessory nucleic acid vector, and/or a nucleic acid vector containing a heterologous nucleic acid. 
     
     
         20 . (canceled) 
     
     
         21 . The method of  claim 16 , wherein the viral vector is selected from a retroviral vector, an adenoviral vector and an adeno-associated viral vector. 
     
     
         22 . The method of  claim 21 , wherein the retroviral vector is a lentiviral vector.